DESCRIPTION: Members of the genus Actinomyces are among a limited number of Gram-positive bacteria which colonize the tooth surface during the early stages of plaque formation. These organisms interact with other plaque bacteria, including various Gram-negative anaerobic species that are found in mature plaques, and often are associated with host tissue inflammation and destruction. Thus, the early tooth colonizers not only play a pivotal role in the initiation of plaque development, but also significantly affect microbial ecology of the oral environment. The Actinomyces fimbriae, designated type 1, have been implicated in mediating adherence of these organisms to the tooth surface. Although type 1 fimbria are thought to bind to salivary acidic, proline-rich protein receptors coated on the tooth surface, it is not known whether additional fimbrial components are involved in this interaction. Moreover, the genes needed for synthesis, assembly, and function of the Actinomyces fimbria, and the mechanisms that regulate fimbrial expression in these organisms, are not known. It has been difficult to design studies to elucidate the molecular nature of fimbriae-mediated adherence of Actinomyces to host tissues primarily due to the lack of a genetic system in these organisms. The objectives of the present study will be to develop a genetic system for Actinomyces utilizing actinobacteriophages as tools, and to initiate characterization of the fimbrial genes of A. viscosus T14V type 1 fimbriae. The first specific aim is to develop transfection procedures for genetic transfer among strains of Actinomyces spp. The second is to characterize three broad- host-range temperate actinobacteriophages and to construct shuttle vectors that replicate in Actinomyces and E. coli. The third is to use shuttle vectors to clone selectable markers/genes and to examine their expression in Actinomyces. The fourth specific aim is to map a region of A. viscosus T14V chromosomal DNA that contains the genetic determinants involved in synthesis and function of fimbriae and to begin characterization of the fimbrial-associated genes. The results of these studies should provide powerful tools and experimental systems for studies of the biological processes involved in fimbriae synthesis and transport in these Gram-positive bacteria. The genetic system will facilitate studies of the functions of various fimbrial genes and their contribution to bacterial adherence. The genetic tools will provide the foundation for future studies on the regulation of fimbriae expression in these organisms in these organisms in their natural ecological niche.